Compositions and methods for reducing body fat

ABSTRACT

The present invention relates to compositions, such as bimatoprost, latanoprost and travoprost, and methods to reduce fat in the body of an individual, for example, by topical administration, injection, and/or implantation of such compositions.

CROSS-REFERENCE TO RELATED APPLICATIONS

The present application is a continuation of and claims priority under35 U.S.C. §120 to U.S. application, U.S. Ser. No. 12/652,968, filed Jan.6, 2010, which is a continuation of and claims priority under 35 U.S.C.§120 to U.S. application, U.S. Ser. No. 11/712,839, filed Mar. 1, 2007,which claims the benefit of and priority to U.S. Provisional PatentApplication, U.S. Ser. No. 60/785,360, filed Mar. 23, 2006, and U.S.Provisional Patent Application, U.S. Ser. No. 60/844,337, filed Sep. 13,2006, the entire disclosures of which are incorporated by referenceherein for all purposes.

FIELD OF THE INVENTION

The present invention relates generally to compounds and methods forreducing fat in the body of an individual. More specifically, body fatmay be reduced by administering compounds identified as cyclopentaneheptanoic acid, 2-cycloalkyl or arylalkyl compounds and derivativesthereof.

BACKGROUND

Excess body fat, which may be diffuse or concentrated on particularparts of the body, plays an important role in a wide array of humandiseases. For many people excess body fat is also a source ofpsychosocial distress and reduced self-esteem. As one form of excessbody fat, obesity is responsible for much of the morbidity and healthcare costs in the United States. It is known to be a risk factor fortype 2 diabetes, hypertension, hyperlipidemia, coronary artery disease,stroke, breast and colon cancer, sleep apnea, gallbladder disease,gastroesophogeal reflux disease, fatty liver, gout, and thromboembolism.Levels of cholesterol, blood pressure, blood sugar and uric acid areusually higher in obese people than in those of normal weight.Overweight people also display increased morbidity from coronary heartdisease. Despite increased awareness of these health risks amongAmericans, the prevalence of obesity in the United States has more thandoubled since the turn of the century. As a result, there has beenconsiderable interest in methods to reduce obesity.

Another problem, which may exist with or without obesity, is excess bodyfat concentrated on particular portion(s) of the body. This may involve,for example, prominent and undesired deposits of fat on the abdomen,buttocks, thighs, arms, and/or chin. Such local accumulations of bodyfat (alternatively known as fat maldistribution) may result fromdisease, hormonal status, or as side effects of medication or othersubstances. Even in the absence of disease, cosmetic considerationsapply to individuals who nevertheless perceive an excess ormaldistribution of fat and wish to have it corrected.

A number of medical conditions are also associated with obesity or localexcesses of body fat. These include Cushing syndrome, pseudo-Cushingsyndrome, drug-induced obesity, HIV-related lipodystrophy,hypothyroidism, pseudohypoparathyroidism, hypothalamic obesity,polycystic ovarian disease, depression, binge eating, Prader-Willisyndrome, Bardet-Biedl syndrome, Cohen syndrome, Down syndrome, Turnersyndrome, growth hormone deficiency, growth hormone resistance, andleptin deficiency or resistance.

Medications known to cause obesity or local excesses of body fat includecortisol and analogs, other corticosteroids, megace, sulfonylureas,antiretrovirals, trycyclic antidepressants, monoamine oxidaseinhibitors, selective serotonin reuptake inhibitors, oralcontraceptives, insulin, risperidone, clozapine, and thiazolidinediones.Some of these medications, such as cortisol and antiretrovirals, cancause profound fat maldistribution with characteristics such as “moonfacies” and a “buffalo hump.”

Changes in hormonal status, including physiologic changes such aspregnancy or menopause, may result in excess body fat. Smoking cessationcommonly leads to weight gain and excess body fat. Trauma may favor theaccumulation of excess body fat by virtue of immobility or disuse of anextremity. Similar problems may affect astronauts or anybody immobilizedfor extended periods of time. Some tumors, most notably lipomas, arecharacterized by local collections of fatty cells that may be amenableto methods used to treat other concentrations of body fat.

A number of diseases that do not involve excess fat may neverthelessbenefit from methods to reduce fat deposits. For example, althoughthyroid orbitopathy (Grave's disease) is not primarily a result ofexcess orbital fat, one treatment is to debulk the orbital fat. Thosewho have suffered hip fractures, for example, show improved outcomeswith weight loss. A number of plastic surgery procedures would benefitfrom adjunctive methods to reduce fat deposits.

Even in the absence of underlying pathology, an individual may havecosmetic concerns about local or diffuse deposits of body fat. These canusually be attributed to constitutional or hereditary factors,developmental history, age, gender, diet, alcohol use, or othercomponents of lifestyle. Individuals in such circumstances commonly wishto reduce the amount of fat on the abdomen, chest, buttocks, hips,thighs, legs, knees, arms, chin, and/or neck. Others may wish to modifyfacial or orbital fat, as in somebody with prominent facial fat pads ororbital fat prolapse.

A number of methods have been developed to reduce or remove excess bodyfat. It is helpful to classify these methods as either extractive ormetabolic. Extractive methods, such as lipoplasty (liposuction) or localexcision, are methods whereby fat is mechanically removed from areas ofinterest. Such methods enable one to focus on particular parts of thebody; however, they are costly and may involve scars, postsurgicaldeformity or regression, discomfort, and adverse reactions.

In contrast to extractive methods, metabolic methods, which includemedications, nutritional supplements, devices, and exercise or otherbody treatment, seek to modify the body's metabolism (whether caloricconsumption, expenditure, or both) such that the body incurs a net lossof fat. Such methods reduce overall body fat but are not particularlysuitable for those who wish to remove body fat from particular areas ofthe body. Another disadvantage is potential concomitant loss of water,carbohydrates, protein, vitamins, minerals, and/or other nutrients.Furthermore, when such methods involve medications, they tend to affectthe entire body. Such medications may have undesired side effects, forexample palpitations, tremor, insomnia, and irritability in those whouse stimulants as appetite suppressants. Despite obvious salubriousvalue, the traditional metabolic methods of diet and exercise are notpractical for everybody.

Therefore, there is a need for new methods and compositions for reducingthe body fat of individuals.

SUMMARY OF THE INVENTION

The present invention describes the use of certain cyclopentaneheptanoic acid, 2-cycloalkyl, or arylalkyl compounds and derivativesthereof, such as bimatoprost, to reduce, or eliminate altogether, bodyfat, for example, adipose tissue in a mammal, for example, a human.These compounds have been described previously, for example in U.S. Pat.Nos. 5,688,819 and 6,403,649. However, it has now been discovered thatthese compounds can be used to reduce body fat.

Accordingly, in one aspect the invention provides a method for reducingfat in a body of an individual, the method including administering tothe skin of a body, injecting into a fat deposit of a body, and/orimplanting an implantable depot into a body, in an amount sufficient toreduce fat in the body, a compound of formula I

in which the dashed bonds represent a single or double bond which can bein the cis or trans configuration; A is alkylene or alkenylene radicalhaving from two to six carbon atoms, which radical may be interrupted byone or more oxide radicals and substituted with one or more hydroxy,oxo, alkyloxy or alkylcarboxy groups, in which the alkyl radical can befrom one to six carbon atoms; B is a cycloalkyl radical having fromthree to seven carbon atoms or an aryl radical selected from the groupconsisting of hydrocarbyl aryl and heteroaryl radicals having from fourto ten carbon atoms in which a heteroatom may be selected from the groupconsisting of nitrogen, oxygen and sulfur atoms; X is a radical selectedfrom the group consisting of —OR⁴ and —N(R⁴)₂ in which R⁴ is selectedfrom the group consisting of hydrogen, a lower alkyl radical having fromone to six carbon atoms, and

in which R⁵ is a lower alkyl radical having from one to six carbonatoms; Z is ═O or represents 2 hydrogen radicals; one of R₁ and R₂ is═O, —OH, or a —O(CO)R₆ group and the other one is —OH or —O(CO)R₆, or R₁is ═O and R₂ is H, in which R₆ is a saturated or unsaturated acyclichydrocarbon group having from 1 to about 20 carbon atoms or —(CH₂)_(m)R₇in which m is 0-10 and R₇ is cycloalkyl radical having from three toseven carbon atoms, or a hydrocarbyl aryl or heteroaryl radical, asdefined above, or a pharmaceutically-acceptable salt thereof, optionallyprovided however that when B is not substituted with a pendantheteroatom-containing radical and Z is ═O, then X is not —OR⁴.

In certain embodiments, the invention provides a method for reducing fatin a body of an individual, the method including administering to theskin of a body, injecting into a fat deposit of a body, and/orimplanting into a body an implantable depot in an amount sufficient toreduce fat in the body, a compound represented by formula II

in which y is 0 or 1, x is 0 or 1 and x and y are not both 1; Y is aradical selected from the group consisting of alkyl, halo, nitro, amino,thiol, hydroxy, alkyloxy, alkylcarboxy and halosubstituted alkyl, inwhich the alkyl radical can be from one to six carbon atoms, n is 0 oran integer of from 1 to 3 and R₃ is ═O, —OH or —O(CO)R₆, andpharmaceutically acceptable salts thereof. In certain circumstances,formula II can be considered an embodiment of formula I.

In certain embodiments, the invention provides a method for reducing fatin a body of an individual, the method including administering to theskin of a body, injecting into a fat deposit of a body, and/orimplanting into a body an implantable depot in an amount sufficient toreduce fat in the body, a compound represented by formula III

in which hatched lines indicate the α configuration and solid trianglesindicate the β configuration, and pharmaceutically acceptable saltsthereof. In certain circumstances, formula III can be considered anembodiment of formula II.

In certain embodiments, the invention provides a method for reducing fatin a body of an individual, the method including administering to theskin of a body, injecting into a fat deposit of a body, and/orimplanting into a body an implantable depot in an amount sufficient toreduce fat in the body, a compound represented by formula IV

and pharmaceutically acceptable salts thereof. In certain circumstances,formula IV can be considered an embodiment of formula III.

In certain embodiments, the invention provides a method for reducing fatin a body of an individual, the method including administering to theskin of a body, injecting into a fat deposit of a body, and/orimplanting into a body an implantable depot in an amount sufficient toreduce fat in the body, a compound represented by formula VII

and pharmaceutically acceptable salts thereof. In certain circumstances,formula VII can be considered an embodiment of formula III.

In certain embodiments, the invention provides a method for reducing fatin a body of an individual, the method including administering to theskin of a body, injecting into a fat deposit of a body, and/orimplanting into a body an implantable depot in an amount sufficient toreduce fat in the body, a compound represented by formula VIII

and pharmaceutically acceptable salts thereof. In certain circumstances,formula VIII can be considered an embodiment of formula III.

In certain embodiments, the invention provides a method for reducing fatin a body of an individual, the method including administering to theskin of a body, injecting into a fat deposit of a body, and/orimplanting into a body an implantable depot in an amount sufficient toreduce fat in the body, a compound represented by formula V

in which hatched lines indicate the α configuration and solid trianglesindicate the β configuration, and in which Y¹ is Cl or trifluoromethyl,and pharmaceutically acceptable salts thereof. In certain circumstances,formula V can be considered an embodiment of formula I.

In certain embodiments, the invention provides a method for reducing fatin a body of an individual, the method including administering to theskin of a body, injecting into a fat deposit of a body, and/orimplanting into a body an implantable depot in an amount sufficient toreduce fat in the body, a compound represented by formula VI

in which Y¹ is Cl or trifluoromethyl, and the position 9- and/or 11-and/or 15-esters of the compound, and pharmaceutically acceptable saltsthereof. This is embodiment can have the following features. Z can be ═Oand X can be NH₂ or an amido radical. X can be selected from the groupconsisting of NH₂ and OCH₃. In certain circumstances, formula VI can beconsidered an embodiment of formula I.

In certain embodiments, the compound can be cyclopentaneheptenol-5-cis-2-(3α-hydroxy-5-phenyl-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentaneheptenamide-5-cis-2-(3α-hydroxy-5-phenyl-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentaneN,N-dimethylheptenamide-5-cis-2-(3α-hydroxy-5-phenyl-1-trans-pentenyl)-3,5-dihydroxy,[1_(α), 2_(β),3_(α),5_(α)]; cyclopentane heptenylmethoxide-5-cis-2-(3α-hydroxy-5-phenyl-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentane heptenylethoxide-5-cis-2-(3α-hydroxy-4-meta-chlorophenoxy-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentaneheptenylamide-5-cis-2-(3α-hydroxy-4-meta-chlorophenoxy-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentaneheptenylamide-5-cis-2-(3α-hydroxy-4-trifluoromethylphenoxy-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentane N-isopropylheptenamide-5-cis-2-(3α-hydroxy-5-phenyl-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentane N-methylheptenamide-5-cis-2-(3α-hydroxy-5-phenyl-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentaneheptenol-5-cis-2-(3α-hydroxy-4-meta-chlorophenoxy-1-trans-butenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentaneheptenamide-5-cis-2-(3α-hydroxy-4-meta-chlorophenoxy-1-trans-butenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; and/or cyclopentaneheptenol-5-cis-2-(3α-hydroxy-5-phenyl-1-trans-pentenyl)3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)].

Further features of any of the above-identified aspects and/orembodiments include the following. Fat reduction can occur at alocalized area, in which the localized area can include the abdomen,chest, buttocks, hips, thighs, legs, knees, arms, chin, neck, face, andcombinations thereof. The individual can have excess body fat as a sideeffect of medication, in which the medication can include acorticosteroid, an antidepressant, and/or a form of insulin. Theindividual can be suffering from a condition such as excess body weight,obesity, fat maldistribution syndrome, HIV, a genetic disordercharacterized at least in part by excess body fat, Grave's disease,diabetes mellitus, present or recent smoking cessation, and combinationsthereof. A method can further include performing a cosmetic or surgicalprocedure, in which the procedure can include mammoplasty,blepharoplasty, abdominoplasty, lipoplasty, liposuction, rhinoplasty,botulinum toxin administration, transverse rectus abdominis muscle flap,orbital decompression, midface lift, sub-orbicularis oculi fat lift,brow lift, and combinations thereof.

For methods including administering one or more of the compounds to theskin of a body, the compound can be delivered in forms including anointment, a lotion, a cream, a patch, a transdermal system, orcombinations thereof. For methods including administering one or more ofthe compounds by injection, the compound can include a subcutaneousinjection, an intramuscular injection, an intralesional injection, orcombinations thereof. For any of the methods described above, one ormore compounds can be delivered in a sustained-release formulation.

In any aspect and/or embodiment of the invention, the individual can bea mammal and the mammal can be a human. Fat reduction can includereducing fat as measured by at least one of volume, size, mass, bulk,density, amount, and/or quantity. Local and/or total fat reduction canbe greater than or equal to 50 percent, greater than or equal to 25percent, greater than or equal to 10 percent, or greater than or equalto 5 percent. Fat reduction can include reducing fat cell amount (forexample, fat cell number), reducing fat cell volume, reducing fat cellmaturation, and/or dedifferentiating a fat cell.

The foregoing aspects and embodiments of the invention may be more fullyunderstood by reference to the following detailed description andclaims.

DETAILED DESCRIPTION OF THE INVENTION

The present invention describes novel uses of certain cyclopentaneheptanoic acid, 2-cycloalkyl or arylalkyl compounds and derivativesthereof, such as bimatoprost, to reduce, or eliminate altogether, bodyfat, for example, adipose tissue, in a mammal, for example, a human.Previously, these compounds were recognized as hypotensive agents (forexample, U.S. Pat. Nos. 5,688,819 and 6,403,649). Specifically, thesecompounds were shown to effect vasodilation and thereby were predictedto relieve symptoms of various diseases associated with increasedpressure, including acute myocardial infarction, vascular thrombosis,hypertension, pulmonary hypertension, ischemic heart disease, congestiveheat failure, and angina pectoris. Specifically, these compounds wereshown to be effective ocular hypotensive agents useful for the treatmentof various ocular hypertensive conditions, including post-surgicaltrabeculectomy ocular hypertensive episodes, post laser trabeculoplastyocular hypertensive episodes, and glaucoma. Glaucoma is a disease of theeye characterized by optic nerve degeneration, most commonly induced byelevated intraocular pressure.

Previous studies have shown that the use of bimatoprost eyedrops for thetreatment of glaucoma does decrease intraocular pressure, but it alsoinduces unwanted side-effects including periocular pigmentary changesinvolving the eyelid and deepening of the lid sulcus of the eye thatreceives the bimatoprost eye drops. (Peplinski et al. (2004) OPTOM. VIS.SCI. 81:574-577). It was previously theorized that the unwantedside-effect of lid sulcus deepening as a function of topical bimatoprostadministration to the eye was caused by a change to Mueller's muscle,however, it was recognized that further investigation would be needed tounderstand the cause of this unwanted side-effect. (Peplinski (2004)).

The present invention involves the recognition that the unwantedside-effect of lid sulcus deepening as a function of topical bimatoprostadministration to the eye is caused by a decease in fat surrounding theeye. Because of this recognition, the present invention extends the useof one or more cyclopentane heptanoic acid, 2-cycloalkyl or arylalkylcompounds and derivatives thereof, including bimatoprost, to reduce fatin body of an individual. An individual may use one or more of thesecompounds to reduce body fat from a particular part of the body, andthis may be accomplished without systemic administration and withminimal if any side effects. These compounds may be administered by anindividual to himself or herself repeatedly and without specialequipment or training, although a medical professional also canadminister such compounds. Daily administration may be adequate (but notnecessarily preferable) to achieve the desired effect, such that theschedule of administration would be convenient.

Without being bound by theory, reduction in fat as a function ofadministration of bimatoprost and/or or one or more other cyclopentaneheptanoic acid, 2-arylalkyl or cycloalkyl compounds or derivativesthereof may include reducing the number of fat cells, reducing thevolume of one or more fat cells, reducing maturation of one or more fatcells, and/or dedifferentiating one or more fat cells. Such effects maybe mediated through prostaglandin or prostaglandin-like receptors, andcompounds according to the invention may be agonists of such receptors.Cyclopentane heptanoic acid, 2-arylalkyl or cycloalkyl compounds andderivatives thereof, such as bimatoprost, are considered to be membersof the class of prostaglandin F2-alpha receptor agonists, which areknown to be inhibitors of adipocyte differentiation and survival(Serrero et al. (1992) BIOCHEM. BIOPHYS. RES. COMMUN. 183:438-442; Lepaket al. (1993) PROSTAGLANDINS 46:511-517; Serrero et al. (1995) BIOCHEM.BIOPHYS. RES. COMMUN. 212:1125-1132; Lepak et al. (1995) ENDOCRINOLOGY136:3222-3229). Accordingly, the fat-reducing properties of thesecompounds, for example, bimatoprost, may relate to its agonism ofprostaglandin or prostaglandin-like receptors. It is contemplated thatsome of these compounds or derivatives, for example, bimatoprost, maypossess physical or pharmacologic properties that render them moresuitable for reducing body fat. The present invention is intended toencompass all of the compounds and derivatives described in U.S. Pat.Nos. 5,688,819 and 6,403,649, as well as any and all analogs that can bereadily synthesized by one skilled in the art.

The present invention therefore envisions new uses of one or morecyclopentane heptanoic acid, 2-arylalkyl or cycloalkyl compounds orderivatives thereof, such as bimatoprost, as local, topical,transdermal, or intralesional therapy to reduce (or in the extreme,eliminate) deposits of body fat. As such, one or more of these compoundsis particularly suitable as a topical, local, or transdermal therapy forany of a number of medical or cosmetic conditions that involve: (1)local excess(es) of body fat; (2) diffuse excess(es) or body fat; (3)maldistribution of body fat; and/or (4) obesity. The invention includesmethods of using one or more of these compounds to reduce body fat. Thepresent invention also includes methods of administering one or more ofthese compounds locally, topically, transdermally, or intralesionallyfor this purpose. Furthermore, the invention envisions administration ofone of more of these compounds to reduce body fat while minimizing oreliminating any significant systemic or local side effects. Numerouspharmaceutical preparations are described that are suitable forapplication or injection to the skin, mucosa, fat deposits, muscle, orsubcutaneous, intramuscular, subdermal, intradermal, subconjunctival,peribulbar, or retrobulbar tissues or spaces. The invention furtherincludes methods for using one or more of these compounds in combinationwith other methods to reduce body fat.

A. Compounds and Derivatives Relating to the Present Invention

The present invention relates to the use of certain cyclopentaneheptanoic acid, 2-cycloalkyl or arylalkyl compounds and derivativesthereof as therapeutic agents, for example, to reduce (or in theextreme, eliminate) body fat (alternatively known as adipose tissue).These therapeutic agents are represented by compounds having thestructural formula I

as defined above. Certain nonacidic cyclopentane heptanoic acid,2-(phenyl alkyl or phenyloxyalkyl) compounds used in accordance with thepresent invention are encompassed by structural formula VI

and their 9- and/or 11- and/or 15-esters. Furthermore, an embodiment ofthe present invention utilizes bimatoprost, also referred to ascyclopentane N-ethylheptenamide-5-cis-2-(3α-hydroxy-5-phenyl-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)] (see U.S. Pat. No. 6,403,649), or(Z)-7-[(1R,2R,3R,5S)-3,5-Dihydroxy-2-[1E,3S)-3-hydroxy-5-phenyl-1-pentenyl]cyclopentyl]-5-N-ethylheptenamide,the compound represented by formula IV

for which the substituents and symbols are as hereinabove defined.Certain compounds utilized in the present invention are represented byformula II

for which the substituents and symbols are as defined above. Additionalcompounds utilized in the present invention are represented by formulaIII

for which the substituents and the symbols are as defined above. Furtheralternative compounds utilized in the present invention are representedby formula V

In all of the above formulae, as well as in those provided hereinafter,dotted lines on bonds between carbons 5 and 6 (C-5), between carbons 13and 14 (C-13), between carbons 8 and 12 (C-8), and between carbons 10and 11 (C-10) indicate a single or a double bond which can be in the cisor trans configuration. Two solid lines indicate a double bond. Hatchedlines at positions C-9, C-11 and C-15 indicate the α configuration. Ifone were to draw the β configuration, a solid triangular line would beused.

In the compounds used in accordance with the present invention,compounds having the C-9 or C-11 or C-15 substituents in the α or βconfiguration are contemplated. As hereinabove mentioned, in allformulae provided herein broken line attachments to the cyclopentanering indicate substituents in the α configuration. Thickened solid lineattachments to the cyclopentane ring indicate substituents in the βconfiguration. Also, the broken line attachment of the hydroxyl group orother substituent to the C-11 and C-15 carbon atoms signifies the αconfiguration.

For the purpose of this invention, unless further limited, the term“alkyl” refers to alkyl groups having from one to ten carbon atoms. Theterm “cycloalkyl” refers to cycloalkyl groups having from three to sevencarbon atoms. The term “aryl” refers to both substituted andunsubstituted aryl groups having from four to ten carbon atoms. The term“saturated or unsaturated acyclic hydrocarbon group” is used to refer tostraight or branched chain, saturated or unsaturated hydrocarbon groupshaving from one to about 6, alternatively one to about 4, carbon atoms.Such groups include alkyl, alkenyl and alkynyl groups of appropriatelengths. In certain embodiments, such groups include an alkyl, forexample, methyl, ethyl, propyl, butyl, pentyl, or hexyl, or an isomericform thereof.

The definition of R₆ may include a cyclic component, —(CH₂)_(m)R₇, inwhich m is 0 or an integer of from 1 to 10, R₇ is an aliphatic ring fromabout 3 to about 7 carbon atoms, or an aromatic or heteroaromatic ring.The “aliphatic ring” may be saturated or unsaturated, for example, asaturated ring having 3-7 carbon atoms, inclusive. As an aromatic ring,R₇ can be phenyl, and the heteroaromatic rings can have oxygen, nitrogenor sulfur as a heteroatom, i.e. R₇ may be thienyl, furanyl, pyridyl,etc. In certain embodiments, m is 0 or an integer of from 1 to 4.

Z is ═O or represents two hydrogen atoms.

X may be selected from the group consisting of —OR⁴ and —N(R⁴)₂ in whichR⁴ is selected from the group consisting of hydrogen, a lower alkylradical having from one to six carbon atoms, and

in which R⁵ is a lower alkyl radical having from one to six carbonatoms.

A compound within the scope of the present invention is bimatoprost,shown as structural formula IV

and pharmaceutically acceptable salts thereof. Bimatoprost is an exampleof a prostamide, and prostamides can be useful according to the presentinvention. The present invention also relates to modified forms ofbimatoprost, for example, bimatoprost with its amide group removed.

Another compound within the scope of the present invention islatanoprost, shown as structural formula VII

and pharmaceutically acceptable salts thereof. Latanoprost is an exampleof a compound expected to have utility and effects on fat similar tothat described for bimatoprost and other compounds disclosed herein.

Another compound within the scope of the present invention istravoprost, shown as structural formula VIII

and pharmaceutically acceptable salts thereof. Travoprost is an exampleof a compound expected to have utility and effects on fat similar tothat described for bimatoprost and other compounds disclosed herein.

Alternative compounds within the scope of the present invention are thecompounds of formula V

in which X is —OH, i.e. cyclopentane heptanoic acid,5-cis-2-(3α-hydroxy-4-m-chlorophenoxy-1-trans-butenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)] and cyclopentanemethylheptenoate-5-cis-2(3α-hydroxy-4-m-chlorophenoxy-1-trans-butenyl)-3,5dihydroxy, [1_(α),2_(β),3_(α),5_(α)] and the 9- and/or 11- and/or15-esters of this compound. (The numbered designations in brackets referto the positions on the cyclopentane ring.)

In addition, the following compounds may be used in the pharmaceuticalcompositions and the methods of treatment of the present invention:cyclopentaneheptenol-5-cis-2-(3α-hydroxy-5-phenyl-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentaneheptenamide-5-cis-2-(3α-hydroxy-5-phenyl-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentaneN,N-dimethylheptenamide-5-cis-2-(3α-hydroxy-5-phenyl-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentane heptenylmethoxide-5-cis-2-(3α-hydroxy-5-phenyl-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentane heptenylethoxide-5-cis-2-(3α-hydroxy-4-meta-chlorophenoxy-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentaneheptenylamide-5-cis-2-(3α-hydroxy-4-meta-chlorophenoxy-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentaneheptenylamide-5-cis-2-(3α-hydroxy-4-trifluoromethylphenoxy-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentane N-isopropylheptenamide-5-cis-2-(3α-hydroxy-5-phenyl-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentane N-methylheptenamide-5-cis-2-(3α-hydroxy-5-phenyl-1-trans-pentenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentaneheptenol-5-cis-2-(3α-hydroxy-4-meta-chlorophenoxy-1-trans-butenyl)-3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)]; cyclopentaneheptenamide-5-cis-2-(3α-hydroxy-4-meta-chlorophenoxy-1-trans-butenyl)-3,5-dihydroxy,[1_(β),2_(β),3_(α),5 _(α)]; and cyclopentaneheptenol-5-cis-2-(3α-hydroxy-5-phenyl-1-trans-pentenyl)3,5-dihydroxy,[1_(α),2_(β),3_(α),5_(α)].

B. Pharmaceutical Formulations

In certain embodiments, one or more cyclopentane heptanoic acid,2-cycloalkyl or arylalkyl compounds or derivatives thereof areincorporated into a pharmaceutical formulation containing apharmaceutically acceptable carrier that is generally suited to local,topical, transdermal, or intralesional drug administration and includingany such material known in the art.

“Therapeutically effective amount” means the level, amount orconcentration of an agent (i.e., an active pharmaceutical ingredient,such as bimatoprost) needed to treat a disease, disorder or conditionwithout causing significant negative or adverse side effects to thetreated tissue.

“Pharmaceutically acceptable salt” is any salt which retains theactivity of the parent compound and does not impart any deleterious orundesirable effect on the subject to whom it is administered and in thecontext in which it is administered. Such salts are those formed withpharmaceutically acceptable cations, e.g., alkali metals, alkali earthmetals, etc.

“Local administration” or “locally administering” means administration(i.e. by injection, implantation, or topical application) by anon-systemic route (such as by a topical, subcutaneous, intramuscular,subdermal, intradermal, transdermal, intralesional, or ophthalmic route,whereby insignificant amounts of the pharmaceutical agent appearsystemically) of a pharmaceutical agent to or to the vicinity of atarget dermal, subdermal, or intralesional area (such as subcutaneousfat) of a patient. For the purpose of the present invention, “injection”means administration by subcutaneous, intramuscular, subdermal,intradermal, intralesional, subconjunctival, peribulbar, or retrobulbarroutes.

Pharmaceutical compositions may be prepared by combining atherapeutically effective amount of at least one compound according tothe present invention, or a pharmaceutically acceptable salt thereof, asan active ingredient, with conventional topically or locally acceptablepharmaceutical excipients, and by preparation of unit dosage formssuitable for topical or local use. These formulations are describedbelow. The therapeutically effective amount typically is between about0.0001 and about 5% (w/v), for example, about 0.001 to about 1.0% (w/v)in liquid formulations. The actual dose of the active compounds of thepresent invention depends on the specific compound, and on the conditionto be treated.

Suitable carriers for the compounds according to the invention areavailable, and the selection of the carrier will depend upon the form ofthe intended pharmaceutical formulation, e.g., as a solution, ointment,lotion, cream, foam, microemulsion, gel, oil, spray, salve, or the like,and may include naturally occurring and/or synthetic materials. It isunderstood that the selected carrier should not adversely affect thecyclopentane heptanoic acid, 2-cycloalkyl or arylalkyl compound orderivative thereof, or other components of the pharmaceuticalformulation.

Suitable carriers for these types of formulations include, but are notlimited to, water, saline, or other aqueous or nonaqueous solutions orsuspensions. Other preferred carriers include Shephard's™ Cream,Aquaphor™, and Cetaphil™ lotion. Other preferred carriers includeointment bases, e.g., polyethylene glycol-1000 (PEG-1000), conventionalcreams such as HEB cream, gels, as well as petroleum jelly and the like.Examples of suitable carriers for use herein include water, alcohols andother nontoxic organic solvents, glycerin, mineral oil, silicone,petroleum jelly, lanolin, fatty acids, vegetable oils, parabens, waxes,and the like. Particularly preferred formulations herein are colorless,odorless solutions, ointments, lotions, creams, microemulsions and gels.

Ointments are semisolid preparations that are typically based onpetrolatum or other petroleum derivatives. The specific ointment base tobe used, as will be appreciated by those skilled in the art, is one thatwill provide for optimum drug delivery, and will provide for otherdesired characteristics as well, e.g., emolliency or the like. As withother carriers or vehicles, an ointment base should be inert, stable,nonirritating and nonsensitizing. As explained in Remington'sPharmaceutical Sciences, 20th ed. (Easton, Pa.: Mack Publishing Company,2000), ointment bases may be grouped in four classes: oleaginous bases;emulsifiable bases; emulsion bases; and water-soluble bases. Oleaginousointment bases include, for example, vegetable oils, fats obtained fromanimals, and semisolid hydrocarbons obtained from petroleum.Emulsifiable ointment bases, also known as absorbent ointment bases,contain little or no water and include, for example, hydroxystearinsulfate, anhydrous lanolin, and hydrophilic petrolatum. Emulsionointment bases are either water-in-oil (W/O) emulsions or oil-in-water(O/W) emulsions, and include, for example, cetyl alcohol, glycerylmonostearate, lanolin, and stearic acid. Preferred water-solubleointment bases are prepared from polyethylene glycols (PEGs) of varyingmolecular weight; again, reference may be had to Remington's, supra, forfurther information.

Lotions are preparations to be applied to the skin surface of anindividual, for example a mammal, for example, a human, withoutfriction, and are typically liquid or semiliquid preparations in whichsolid particles, including the active agent, are present in a water oralcohol base. Lotions are usually suspensions of solids and can includea liquid oily emulsion of the oil-in-water type. Lotions are preferredformulations herein for treating large body areas, because of the easeof applying a more fluid composition. It is generally necessary that theinsoluble matter in a lotion be finely divided. Lotions will typicallycontain suspending agents to produce better dispersions as well ascompounds useful for localizing and holding the active agent in contactwith the skin, e.g., methylcellulose, sodium carboxymethylcellulose, orthe like. Certain formulations for use in conjunction with the presentinvention contain propylene glycol mixed with a hydrophilic petrolatumsuch as that which may be obtained under the trademark Aquaphor™ fromBeiersdorf, Inc. (Norwalk, Conn.).

Creams containing the active agent can be viscous liquid or semisolidemulsions, either oil-in-water or water-in-oil. Cream bases arewater-washable, and contain an oil phase, an emulsifier, and an aqueousphase. The oil phase is generally includes petrolatum and a fattyalcohol such as cetyl or stearyl alcohol; the aqueous phase usually,although not necessarily, exceeds the oil phase in volume, and generallycontains a humectant. The emulsifier in a cream formulation, asexplained in Remington's, supra, is generally a nonionic, anionic,cationic, or amphoteric surfactant.

Microemulsions are thermodynamically stable, isotropically cleardispersions of two immiscible liquids, such as oil and water, stabilizedby an interfacial film of surfactant molecules (Encyclopedia ofPharmaceutical Technology, vol. 9 (1992) New York: Marcel Dekker). Forthe preparation of microemulsions, a surfactant (emulsifier), aco-surfactant (co-emulsifier), an oil phase, and a water phase arenecessary. Suitable surfactants include any surfactants that are usefulin the preparation of emulsions, e.g., emulsifiers that are typicallyused in the preparation of creams. The co-surfactant (or “co-emulsifer”)is generally selected from the group of polyglycerol derivatives,glycerol derivatives, and fatty alcohols. Certainemulsifier/co-emulsifier combinations are generally although notnecessarily selected from the group consisting of: glyceryl monostearateand polyoxyethylene stearate; polyethylene glycol and ethylene glycolpalmitostearate; and caprilic and capric triglycerides and oleoylmacrogolglycerides. The water phase includes not only water but also,typically, buffers, glucose, propylene glycol, polyethylene glycols, forexample, lower molecular weight polyethylene glycols (e.g., PEG 300 andPEG 400), and/or glycerol, and the like, while the oil phase willgenerally include, for example, fatty acid esters, modified vegetableoils, silicone oils, mixtures of mono- di- and triglycerides, mono- anddi-esters of PEG (e.g., oleoyl macrogol glycerides), and so forth.

Gel formulations are semisolid systems including of either smallinorganic particle suspensions (two-phase systems) or large organicmolecules distributed substantially uniformly throughout a carrierliquid (single phase gels). Single phase gels can be made, for example,by combining the active agent, a carrier liquid and a suitable gellingagent such as tragacanth (at 2 to 5%), sodium alginate (at 2-10%),gelatin (at 2-15%), methylcellulose (at 3-5%), sodiumcarboxymethylcellulose (at 2-5%), carbomer (at 0.3-5%) or polyvinylalcohol (at 10-20%) together and mixing until a characteristic semisolidproduct is produced. Other suitable gelling agents includemethylhydroxycellulose, polyoxyethylene-polyoxypropylene,hydroxyethylcellulose and gelatin. Although gels commonly employ aqueouscarrier liquid, alcohols and oils can be used as the carrier liquid aswell.

A variety of sustained-release formulations, such as patches, pills, orimplantable depots, can used in connection with the present inventionand readily made by one skilled in the art.

Various additives may be included in the topical formulations of theinvention. Examples of additives include, but are not limited to,solubilizers, skin permeation enhancers, opacifiers, preservatives(e.g., anti-oxidants), gelling agents, buffering agents, surfactants(particularly nonionic and amphoteric surfactants), emulsifiers,emollients, thickening agents, stabilizers, humectants, colorants,fragrance, and the like. Inclusion of solubilizers and/or skinpermeation enhancers is particularly preferred, along with emulsifiers,emollients, and preservatives.

Examples of solubilizers include, but are not limited to, the following:hydrophilic ethers such as diethylene glycol monoethyl ether(ethoxydiglycol, available commercially as Transcutol™) and diethyleneglycol monoethyl ether oleate (available commercially as Softcutol™);polyethylene castor oil derivatives such as polyoxy 35 castor oil,polyoxy 40 hydrogenated castor oil, etc.; polyethylene glycol,particularly lower molecular weight polyethylene glycols such as PEG 300and PEG 400, and polyethylene glycol derivatives such as PEG-8caprylic/capric glycerides (available commercially as Labrasol™); alkylmethyl sulfoxides such as DMSO; pyrrolidones such as 2-pyrrolidone andN-methyl-2-pyrrolidone; and DMA. Many solubilizers can also act asabsorption enhancers. A single solubilizer may be incorporated into theformulation, or a mixture of solubilizers may be incorporated therein.

Suitable emulsifiers and co-emulsifiers include, without limitation,those emulsifiers and co-emulsifiers described with respect tomicroemulsion formulations. Emollients include, for example, propyleneglycol, glycerol, isopropyl myristate, polypropylene glycol-2 (PPG-2)myristyl ether propionate, and the like.

Other active agents may also be included in the formulation, e.g.,anti-inflammatory agents, analgesics, antimicrobial agents, antifungalagents, antibiotics, vitamins, antioxidants, and sunblock agentscommonly found in sunscreen formulations including, but not limited to,anthranilates, benzophenones (particularly benzophenone-3), camphorderivatives, cinnamates (e.g., octyl methoxycinnamate), dibenzoylmethanes (e.g., butyl methoxydibenzoyl methane), p-aminobenzoic acid(PABA) and derivatives thereof, and salicylates (e.g., octylsalicylate).

Also, the pharmaceutical formulation may be sterilized or mixed withauxiliary agents, e.g., preservatives, stabilizers, wetting agents,buffers, or salts for influencing osmotic pressure and the like. Sterileinjectable solutions can be prepared by incorporating one or morecyclopentane heptanoic acid, 2-cycloalkyl or arylalkyl compounds orderivatives thereof in the required amount in the appropriate solventwith various other ingredients as enumerated above, as required,followed by filtered sterilization. Generally, dispersions are preparedby incorporating the various sterilized active ingredients into asterile vehicle which contains the basic dispersion medium and therequired other ingredients from those enumerated above. In the case ofsterile powders for the preparation of sterile injectable solutions, thepreferred methods of preparation are vacuum-drying and freeze-dryingtechniques which yield a powder of the active ingredient plus anyadditional desired ingredient from a previously sterile-filteredsolution thereof.

For ophthalmic application, solutions typically are prepared using aphysiological saline solution as a major vehicle. The pH of suchophthalmic solutions should be maintained between 4.5 and 8.0 with anappropriate buffer system, a neutral pH being preferred but notessential. The formulations may also contain conventional,pharmaceutically acceptable preservatives, stabilizers and surfactants.

Certain preservatives that may be used for ophthalmic preparation(s)include, but are not limited to, benzalkonium chloride, chlorobutanol,thimerosal, phenylmercuric acetate and phenylmercuric nitrate. Onesurfactant that may be used is, for example, Tween 80. Likewise, variousvehicles may be used in the ophthalmic preparations of the presentinvention. These vehicles include, but are not limited to, polyvinylalcohol, povidone, hydroxypropyl methyl cellulose, poloxamers,carboxymethyl cellulose, hydroxyethyl cellulose cyclodextrin andpurified water. Tonicity adjustors may be added as needed or convenient.They include, but are not limited to, salts, particularly sodiumchloride, potassium chloride, mannitol and glycerin, or any othersuitable ophthalmically acceptable tonicity adjustor. Various buffersand means for adjusting pH may be used so long as the resultingpreparation is ophthalmically acceptable. Accordingly, buffers includeacetate buffers, citrate buffers, phosphate buffers and borate buffers.Acids or bases may be used to adjust the pH of these formulations asneeded. In a similar vein, an ophthalmically acceptable antioxidant foruse in ophthalmic preparation(s) includes, but is not limited to, sodiummetabisulfite, sodium thiosulfate, acetylcysteine, butylatedhydroxyanisole and butylated hydroxytoluene.

C. Uses of the Compositions and Compounds of the Invention

The present invention uses one or more cyclopentane heptanoic acid,2-arylalkyl or cycloalkyl compounds and derivatives thereof to reduce(or in the extreme, eliminate) body fat. For the purpose of thisinvention, unless further limited, the term “reduce” means to diminishthe volume, size, mass, bulk, density, amount, and/or quantity of asubstance. The present invention is expected to reduce fat by greaterthan or equal to 50%, by greater than or equal to 25%, by greater thanor equal to 10%, and/or by greater than or equal to 5%. For example, fatreduction can include reducing fat cell amount (for example, fat cellnumber), reducing fat cell volume, reducing fat cell maturation, and/ordedifferentiating a fat cell.

Specifically, the invention envisions new uses for these compounds whenadministered locally or topically to the skin, fat deposits, muscle,subcutaneous space, intramuscular space, eye, orbital tissue, orbitalspace(s), tumor, lesion, or mucous membrane of an individual, forexample, a mammal, for example, a human. The invention contemplatesspecial usefulness for individuals with obesity, fat maldistribution, orcosmetic disturbances of excess or maldistributed body fat, whetherlocal or diffuse.

The invention is further expected to be useful for individuals withprominent or undesired deposits of fat on the abdomen, chest, buttocks,hips, thighs, legs, knees, arms, chin, face, or neck. The invention isalso conceived to be beneficial for individuals with Cushing syndrome,pseudo-Cushing syndrome, drug-induced obesity, HIV-relatedlipodystrophy, hypothyroidism, pseudohypoparathyroidism, hypothalamicobesity, polycystic ovarian disease, depression, binge eating,Prader-Willi syndrome, Bardet-Biedl syndrome, Cohen syndrome, Downsyndrome, Turner syndrome, growth hormone deficiency, growth hormoneresistance, or leptin deficiency or resistance. The invention is furthercontemplated to be of utility for individuals using cortisol andanalogs, other corticosteroids, megace, sulfonylureas, trycyclicantidepressants, monoamine oxidase inhibitors, selective serotoninreuptake inhibitors, oral contraceptives, insulin, risperidone,clozapine, and thiazolidinediones.

It is of note that corticosteroids, which cause inappropriate localaccumulations of fat on various body parts (Cushingoid appearance), arepotent inhibitors of prostaglandin pathways, and that thisantiprostaglandin activity may mediate the Cushingoid appearance. Thecompounds and derivatives of the present invention may therefore beuseful as specific, local opponents to this process in individuals onsystemic corticosteroid therapy.

The invention is also conceived to be useful for individuals with anyhormonal status, including physiologic changes such as pregnancy ormenopause, that results in excess body fat. The invention is furtherenvisioned to be beneficial for treating weight gain or excess body fatin individuals who are undergoing or who have recently undergone smokingcessation. Further uses of the invention include treatment ofindividuals for whom immobility or disuse of an extremity favors theaccumulation of excess body fat. The invention is also contemplated tobe useful for individuals with thyroid orbitopathy (Grave's disease),whereby reduction of orbital fat may lead to improvement ofexophthalmos, eyelid displacement, signs and symptoms or ocularexposure, or optic neuropathy. It is further envisioned that theinvention will be useful for achieving weight loss in individuals whohave suffered hip fractures. Likewise, the invention is expected to beuseful for astronauts and others who face prolonged periods ofimmobility.

The invention may also be beneficial in the treatment of fatty tumorssuch as lipomas, regardless of where they occur in the body.

The invention may also be useful as an adjunct to other cosmetic orsurgical procedures, including but not limited, to mammoplasty,blepharoplasty, abdominoplasty, lipoplasty (liposuction), rhinoplasty,botulinum toxin (Botox™) administration, transverse rectus abdominismuscle (TRAM) flap, orbital decompression, midface lift, sub-orbicularisoculi fat (SOOF) lift, and brow lift.

The invention may also be useful as an adjunct to any of various kindsof surgery, whether used in the pre-operative, peri-operative, orpost-operative period. The invention further contemplates uses precedingabdominal, thoracic, oncologic, endocrine, neurologic, transplant, anddermatologic surgery, whereby surgical exposure may be improved; andpreceding or following orthopedic procedures, whereby surgical exposureas well as post-operative recovery may be improved.

Throughout the description, where compositions are described as having,including, or comprising specific components, or where processes aredescribed as having, including, or comprising specific process steps, itis contemplated that compositions of the present invention also consistessentially of, or consist of, the recited components, and that theprocesses of the present invention also consist essentially of, orconsist of, the recited processing steps. Further, it should beunderstood that the order of steps or order for performing certainactions are immaterial so long as the invention remains operable.Moreover, two or more steps or actions may be conducted simultaneously.

EXAMPLES

In light of the foregoing description, the specific non-limitingexamples presented below are for illustrative purposes and not intendedto limit the scope of the invention in any way. In Examples 4-8, variousmodes of non-systemic administration of a cyclopentane heptanoic acid,2-arylalkyl or cycloalkyl compound or derivative thereof can be used,for example, by topical application (ointment, cream, or transdermalpatch), subcutaneous injection, intramuscular injection, orintralesional injection, any of which may involve a controlled (orsustained) release formulation or depot.

Example 1 In Vitro Study of Mouse Preadipocyte Viability, PreadipocyteDifferentiation, Preadipocyte Apoptosis, and Adipocyte Apoptosis

The following experiment describes administration of bimatoprost, anexample of compounds according to the invention, to adipocytes andpreadipocytes to determine cell viability, apoptosis, and celldifferentiation as a function of the presence of bimatoprost and/or as afunction of the concentration of bimatoprost. The protocol is generallyadapted from Lin et al. (2005) OBESITY 13: 982-990.

Preparation of Preadipocytes.

3T3-L1 mouse embryo fibroblasts are obtained and cultured. Cells arecultured at 37° C. in a humidified 5% CO₂ atmosphere and grown inDulbecco's modified Eagle's medium (DMEM) with 10% bovine calf serum.Confluency is induced for 2 days (Days −1 and 0) and preadipocytes arecultured in DMEM with 10% fetal bovine serum (FBS/DMEM) medium,supplemented with 167 nM insulin, 0.5 μM isobutylmethylxanthine, and 1μM dexamethasone for 2 days (Days 1 and 2).

Preparation (Induction) of Mature Adipocytes from Preadipocytes.

Preadipocytes prepared as above are then maintained in a culture mediumwith 167 nM insulin supplement for another 2 days (Days 3 and 4),followed by culturing with 10% FBS/DMEM medium for an additional 4 days(Days 5-8), at which time >90% of the cells are expected to havedifferentiated and matured into adipocytes with accumulated fatdroplets. All media contains 100 U/mL penicillin, 100 μg/mLstreptomycin, and 292 μg of L-glutamine/mL.

All experiments, except where otherwise indicated, are performed withbimatoprost added to the culture medium for a final concentration of 0%,0.003%, 0.03%, or 0.3% for a duration of 3, 6, 12, or 24 hours [with1:1000 dimethyl sulfoxide (DMSO)]. In alternative experimental protocolsaccording to this example, the duration of exposure to bimatoprost is onthe order of days, weeks, or months. To the extent longer exposure timesare desired, certain aspects of the experimental protocols described inthis example are altered accordingly, for example, to maintain cellviability in culture over longer periods of exposure.

Test for Cell Viability.

Standard MTS tests of cell viability are performed in 96-well plates.MTS is a tetrazolium compound that is converted to formazan withinmetabolically active cells. Non-viable cells do not perform thisconversion. For mature adipocytes, seeding density is 5000 cells/well,and cells undergo induction until matured. For preadipocytes, seedingdensity is 2500 cells/well, and cells are cultured overnight beforetreatment (with an expected doubling time of approximately 18 hours).Cells are incubated with either DMSO (1:1000) or increasingconcentrations of bimatoprost for up to 24 hours. The medium is thenchanged and replaced with 100 μL of fresh 10% FBS and 20 μL of MTSsolution (Promega, Madison, Wis.). Cells are then returned to theincubator for an additional 2 hours before 25 μL of 10% sodium dodecylsulfate is added to stop the reaction. The plate is analyzed byspectrometry at a wavelength of 490 nm, which is an absorbance peak forformazan. The quantity of formazan as measured by absorbance at 490 nmis a directly proportional to the number of viable cells.

Test for Apoptosis (1).

Laser scanning cytometry (LSC) is employed to detect apoptosis-mediateddisruption of the plasma membrane. Mature adipocytes are incubated witheither DMSO (1:1000) or increasing concentrations of bimatoprost for 24hours. Monolayer cells from each treatment are washed twice with coldphosphate-buffered saline (PBS) and once in binding buffer and thenincubated in the dark with gentle agitation for 10 minutes with 5 μL ofannexin V-fluorescein isothiocyanate (AV, which bindsphosphatidylserine) and 5 μL of propidium iodide (PI) in 450 μL ofbinding buffer (BD Biosciences, San Diego, Calif.) at ambienttemperature. LSC is carried out as previously described in the art.Apoptotic cells fluoresce and are detected by LSC, since AV bindsspecifically to phosphatidylserine in the disrupted plasma membrane. PI,which binds specifically to double-stranded DNA, is only exposed toextracellular milieu in late apoptosis or cell death. Cells aretherefore characterized as follows: viable (low AV and PI intensity),apoptotic (high AV and low PI intensity), or dead (high PI intensity).

Test for Apoptosis (2).

Terminal Deoxynucleotidyl Transferase dUTP Nick-End Labeling (TUNEL)Imaging is one of the most common assays for apoptosis. The TUNEL methodincorporates labeled terminal nucleotides to detect apoptosis-mediatedcleavage and degradation of DNA. Both preadipocytes and matureadipocytes are incubated with or without bimatoprost for 24 hours. AnAPO-BrdU TUNEL kit can obtained from a supplier. Monolayer cells arefixed with 10% formalin (PBS buffered) and stored in 70% ethanol at −20°C. for 24 hours. Cells are washed twice in the washing buffer, followedby one-hour incubation in a humidity box at 37° C. with gentle agitationin 150 μL of DNA-labeling solution (containing 30 μL of reaction buffer,2.25 μL of terminal deoxynucleotidyl transferase, 24 μL of BrdUTP, and93.75 μL of dH₂O). After a brief wash, the cells are incubated for 30minutes with Alexa Fluor 488 dye-labeled anti-BrdU antibody (1:20dilution), followed by an additional 30 minutes with PI/RNase buffer tostain the nucleus at room temperature in the dark, and viewed with afluorescent microscope. Apoptosis manifests as fluorescence uponmicroscopic examination, which reflects the antibody-labeled BrdUincorporated at the cleaved portions of DNA.

Test for Differentiation of Preadipocytes into Mature Adipocytes.

Oil-Red-O, a specific lipid stain, is used to test the effect ofbimatoprost on adipogenesis. Three final concentrations of bimatoprost(0.003%, 0.03%, and 0.3% μM), along with 1:1000 DMSO control are addedwith the induction medium (see schedule above) in addition tono-addition normal controls. The medium is changed every 2 days. Toidentify cells undergoing adipogenesis, cells are stained withOil-Red-O. Briefly, dishes are washed with cold PBS and fixed in 10%neutral formalin. After two changes of propylene glycol, Oil-Red-0 isadded with agitation for 7 minutes, followed by washing in 85% propyleneglycol. The dishes are then rinsed in distilled water and counterstainedwith hematoxylin. For each dish, multiple images are taken and analyzedfor average lipid droplet size, percentage lipid area, and total dropletnumber with software. Lipid droplets appear as red-stained features.

Results.

Specific observations which are anticipated include: (1) on MTS testing,lower absorbance at 490 nm of bimatoprost-treated cells compared tocontrol cells; (2) on laser scanning cytometry (LSC), more AV-labeledand/or PI-labeled (apoptotic or dead) cells among bimatoprost-treatedsamples compared to control samples; (3) on TUNEL imaging, moreBrdU-antibody-labeled (fluorescent) cells among bimatoprost-treatedsamples compared to control samples; and (4) on Oil-Red-0 staining,smaller and fewer lipid droplets seen on bimatoprost-treated samplescompared to control samples (reflecting a lower degree ofdifferentiation of preadipocytes into adipocytes).

Conclusion.

The experiments described above are designed to and are contemplated toprovide a conclusion that (1) preadipocytes and/or mature adipocytestreated with bimatoprost have lower rates of viability compared tocontrol samples; (2) preadipocytes and/or mature adipocytes treated withbimatoprost have higher rates of apoptosis compared to control samples;and/or (3) preadipocytes treated with bimatoprost have lower rates ofdifferentiation into mature adipocytes compared to control samples.These results may correlate in a dose-dependent manner within a range ofbimatoprost administration. As a result, these contemplated outcomeswill show, in vitro, that compounds according to the present invention,including bimatoprost, can reduce fat in one or more of a variety ofways, including cell death and reduced cell maturation.

Example 2 In Vivo Rodent Study

The following experiment describes a randomized study in laboratoryrodents to test whether compounds according to the invention, includingtopical bimatoprost, reduce total body fat mass, subcutaneous fat mass,and total body mass.

Animal Preparation.

Genetically uniform laboratory rodents (for example, hairless rats ornude mice) are used. They are adult laboratory rodents similar in sizeat the commencement of the study (approximately postnatal week 15 forrats). Multiple animals are tested. Animals randomized to bimatoprostare housed separately from control animals but are kept in the same roomunder identical environmental conditions. All animals are fed a standarddiet, appropriate for the species, ad libidum.

Method.

In all cases the vehicle is a compatible ointment, such as petrolatum,with a final bimatoprost concentration of 0%, 0.003%, 0.03%, or 0.3%. Afixed amount of ointment, for example 1 gram, is applied daily in a thinfilm over a uniform surface area of the abdominal skin of each animal.Each animal is randomized to a daily application of a dose of topicalbimatoprost or vehicle alone. On a weekly basis, total body mass of eachanimal is determined on a scale. Total body fat is estimated by any ofthe several methods established for this purpose, for example, waterdisplacement (Dahms et al. (1982) J NUTR. 112: 398-400), isotopicdilution (Culebras et al. (1977) J. PHYSIOL. 232: R60-65), Total BodyElectrical Conductance (TOBEC) (Baer et al. (1993) PHYSIOL. BEHAV. 53:1195-99), Dual-Energy X-Ray Absorptiometry (DEXA) (Bertin et al. (1998)J NUTRITION 128: 1550-54), or Nuclear Magnetic Resonance (NMR) (Kunneckeet al. (2004) OBES RES. 12: 1604-15). Animals are maintained andmonitored in this fashion for six months, at which point they aresacrificed.

Analysis and Results.

Abdominal fat pads of the sacrificed animals are excised and measureddirectly for mass and volume. Direct carcass analysis with chemicalextraction may be further performed (Frisch et al. (1977) PNAS 74:379-383). All results are normalized with reference to baseline valuesand then compared between bimatoprost and control groups.

Conclusion.

It is contemplated that the experiment described above will indicatethat animals treated with compounds according to the present invention,in this case bimatoprost, will exhibit lower total body fat mass, lowersubcutaneous (abdominal) fat mass, and/or lower total body mass comparedto control animals. These results may correlate in a dose-dependentmanner within a range of bimatoprost administration. As a result, thesecontemplated outcomes will show, in vivo, that compounds according tothe present invention, including bimatoprost, can reduce fat.

Example 3 In Vivo Human Study

The following experiment describes a randomized, double-blind study inhuman subjects to test whether compounds according to the invention,including topical bimatoprost, reduces fat in the body of individuals.

Multiple human subjects (both male and female), for example, with bodymass indices of 30 or more but otherwise healthy, are entered into arandomized double-blind study. Either the left or the right arm israndomized to receive topical bimatoprost; the other arm receivesvehicle only. Bimatoprost is supplied in a petrolatum-based ointment(the vehicle) at a final concentration of 0%, 0.003%, 0.03% or 0.3%.Ointment containers are unlabeled as to the presence or concentration ofbimatoprost.

Each day, subjects apply a thin film of ointment to the skin over therespective triceps while wearing new, clean surgical gloves. Subjectsare instructed to refrain from washing the treated area for at least 8hours and are instructed to refrain from wearing tight clothing orocclusive dressings that will come into contact with the treated area.

Body fat calipers are used to measure fat in the triceps regionbilaterally on a weekly basis. The study continues for 6 months. It iscontemplated that the experiments described above will indicate thatbody areas, such as arms, treated with compounds according to thepresent invention, including bimatoprost, will exhibit lower fatcompared to control areas, such as arms, treated with vehicle alone.These results may correlate in a dose-dependent manner within a range ofbimatoprost administration. As a result, this contemplated outcome willshow, in vivo, that compounds according to the present invention,including bimatoprost, can reduce fat.

Example 4 Use of Cyclopentane Heptanoic Acid, 2-Arylalkyl or CycloalkylCompounds and Derivatives Thereof to Reduce Obesity

The following description exemplifies a clinical application of acyclopentane heptanoic acid, 2-arylalkyl or cycloalkyl compound or aderivative thereof to treat obesity.

A 52-year-old man is diagnosed with type 2 diabetes and hypertension.Medical evaluation reveals that his height is 5 feet 8 inches and aweight of 211 pounds, corresponding to a body mass index (BMI) of 32.1(clinically obese). His noninvasive blood pressure is 184/98. Hisfasting serum glucose is 147 and his hemoglobin A1C is 7.6. His physicalexam is notable for prominent central (abdominal) obesity.

In addition to medical therapy, the man's physician recommends a regimenof diet and exercise. After six months the man is unable to lose weight.The physician prescribes daily topical application of a bimatoprostointment to the abdomen as treatment for central obesity. After a periodof time, for example from a few days to several months, the man's BMI isreduced and/or abdominal fat deposits are reduced.

Example 5 Use of Cyclopentane Heptanoic Acid, 2-Arylalkyl or CycloalkylCompounds and Derivatives Thereof to Treat Signs and Symptoms of CushingDisease

The following description exemplifies a clinical application of acyclopentane heptanoic acid, 2-arylalkyl or cycloalkyl compound or aderivative thereof to treat Cushing disease.

A 19-year-old woman with severe persistent asthma is dependent onhigh-dose prednisone for several months. Although she is able tomaintain normal weight (BMI 21), there are pronounced fatty deposits onher cheeks (so-called “moon facies”) and a fat pad between her shoulders(so-called “buffalo hump”). She is diagnosed with Cushing syndromesecondary to prolonged prednisone therapy. Attempts to taper theprednisone are thwarted by multiple asthma exacerbations requiringhospitalization, including one requiring intubation.

The woman's physician prescribes a daily application of a bimatoprostointment to the face and between the shoulders as treatment for the fatdeposits. After a period of time, for example from a few days to severalmonths, the fatty deposits on the woman's cheeks are reduced and/or thefat pad between her shoulders is reduced.

Example 6 Use of Cyclopentane Heptanoic Acid, 2-Arylalkyl or CycloalkylCompounds and Derivatives Thereof to Treat Fat MaldistributionAssociated with HIV/AIDS

The following description exemplifies a clinical application of acyclopentane heptanoic acid, 2-arylalkyl or cycloalkyl compound or aderivative thereof to treat fat maldistribution associated withHIV/AIDS.

A 34-year-old man with HIV/AIDS uses multiple reverse transcriptaseinhibitors and protease inhibitors for seven years, since first beingdiagnosed as HIV seropositive. Despite normal body weight (BMI 20),there are pronounced fatty deposits on his cheeks (so-called “moonfacies”) and a fat pad between his shoulders (so-called “buffalo hump”).He is diagnosed with HIV-related fat maldistribution syndrome(lipodystrophy).

The man's physician prescribes a daily application of a bimatoprostointment to the face and between the shoulders as treatment for the fatdeposits. After a period of time, for example from a few days to severalmonths, the fatty deposits on the man's cheeks are reduced and/or thefat pad between his shoulders is reduced.

Example 7 Use of Cyclopentane Heptanoic Acid, 2-Arylalkyl or CycloalkylCompounds and Derivatives Thereof to Reduce Local Fat Deposits ofFunctional and/or Cosmetic Significance

The following description exemplifies a clinical application of acyclopentane heptanoic acid, 2-arylalkyl or cycloalkyl compound or aderivative thereof to reduce local fat deposits of functional and/orcosmetic significance.

A 56-year-old female flight attendant is troubled by prominent fatdeposits on her hips and thighs, which interfere with her work and lowerher self-esteem. Her physician recommends diet and exercise. The womanloses 7 pounds, but there is no noticeable reduction in the fatdeposits. She is referred to a plastic surgeon but declines lipoplastydue to potential adverse effects.

The plastic surgeon prescribes a daily application of a bimatoprostointment to the hips and thighs as treatment for the fat deposits. Aftera period of time, for example from a few days to several months, thefatty deposits on the woman's hips and/or thighs are reduced.

Example 8 Use of Cyclopentane Heptanoic Acid, 2-Arylalkyl or CycloalkylCompounds and Derivatives Thereof to Treat the Orbitopathy of Grave'sDisease (Thyroid Orbitopathy)

The following description exemplifies a clinical application of acyclopentane heptanoic acid, 2-arylalkyl or cycloalkyl compound or aderivative thereof to treat the orbitopathy of Grave's disease (thyroidorbitopathy).

A 42-year-old woman with Grave's disease sees an oculoplastic specialistfor complaints of bilateral eye discomfort. Eye exam reveals normalvision and no evidence of compressive optic neuropathy; however, thereis mild bilateral proptosis, lid retraction, and evidence of cornealexposure. The woman wishes to avoid systemic steroids if possible.

The doctor prescribes an ophthalmic preparation of bimatoprost daily toboth eyes as a treatment for the proptosis and exposure symptoms. Aftera period of time, for example from a few days to several months, thewoman's orbital fat is reduced and her condition improves.

INCORPORATION BY REFERENCE

The entire disclosure of each of the publications and patent documentsreferred to herein is incorporated by reference in its entirety for allpurposes to the same extent as if each individual publication or patentdocument were so individually denoted.

EQUIVALENTS

The invention may be embodied in other specific forms without departingfrom the spirit or essential characteristics thereof. The foregoingembodiments are therefore to be considered in all respects illustrativerather than limiting on the invention described herein. Scope of theinvention is thus indicated by the appended claims rather than by theforegoing description, and all changes that come within the meaning andrange of equivalency of the claims are intended to be embraced therein.

What is claimed is:
 1. A method for reducing fat in a body of anindividual, the method comprising administering to the skin of a portionof the body, travoprost, represented by formula VIII:

or a pharmaceutically acceptable salt thereof, wherein the portion ofthe body has an excess deposit of body fat.
 2. The method of claim 1,wherein fat reduction occurs at the portion of the body.
 3. The methodof claim 1, wherein the portion of the body is selected from the groupconsisting of abdomen, chest, buttocks, hips, thighs, legs, knees, arms,chin, neck, face, and combinations thereof.
 4. The method of claim 1,wherein the individual has excess body fat as a side effect ofmedication.
 5. The method of claim 1, wherein the individual suffersfrom a condition selected from the group consisting of excess bodyweight, obesity, fat maldistribution syndrome, HIV, a genetic disordercharacterized at least in part by excess body fat, Grave's disease,diabetes mellitus, present or recent smoking cessation, and combinationsthereof.
 6. The method of claim 1, further comprising performing acosmetic or surgical procedure.
 7. The method of claim 6, wherein theprocedure is selected from the group consisting of mammoplasty,blepharoplasty, abdominoplasty, lipoplasty, liposuction, rhinoplasty,botulinum toxin administration, orbital decompression, midface lift,sub-orbicularis oculi fat lift, brow lift, and combinations thereof. 8.The method of claim 1, wherein said compound is delivered in a formselected from the group consisting of an ointment, a lotion, a cream, apatch, and a transdermal system.
 9. The method of claim 1, wherein thecompound is in a sustained-release formulation.
 10. The method accordingto claim 1, wherein the individual is a human.
 11. The method of claim1, wherein fat reduction comprises reducing fat as measured by at leastone of volume, size, mass, bulk, density, amount, or quantity.
 12. Themethod of claim 11, wherein the reduction of local or total fat isgreater than or equal to 50 percent, greater than or equal to 25percent, greater than or equal to 10 percent, or greater than or equalto 5 percent.
 13. The method of claim 1, wherein fat reduction comprisesreducing fat cell amount.
 14. The method of claim 1, wherein fatreduction comprises reducing fat cell volume.
 15. The method of claim 1,wherein fat reduction comprises reducing fat cell maturation.
 16. Themethod of claim 1, wherein fat reduction comprises dedifferentiating afat cell.
 17. The method of claim 1, wherein the skin is the skin of theabdomen, chest, buttocks, hips, thighs, legs, knees, arms, chin, neck,face, or combinations thereof.
 18. The method of claim 17, wherein theskin is skin on the face.
 19. The method of claim 1, wherein the skin isperiorbital skin.
 20. The method of claim 1, wherein the method reducesorbital fat.
 21. The method of claim 1, wherein the method reducesfacial fat.
 22. The method of claim 1, wherein the individual suffersfrom thyroid orbitopathy.
 23. The method of claim 1, wherein theindividual suffers from Grave's disease.
 24. The method of claim 1,wherein the individual has prominent facial fat pads.
 25. The method ofclaim 1, wherein the individual has orbital fat prolapse.